ripk1 kinase inhibitor necrostatin 1  (MedChemExpress)

Journal: Cell Death & Disease

Article Title: TAK1 inhibition leads to RIPK1-dependent apoptosis in immune-activated cancers

doi: 10.1038/s41419-024-06654-1

Figure Lengend Snippet: A Schematic cartoon of TAK1 depletion using a dTAG-TAK1 degradation system. B Western blots of time course experiment treating dTAG-TAK1 GSCs with 100 nM dTAG V -1 ligand for indicated amount of time. C Barplot of total % Annexin V positive cells quantified by flow cytometry after treatment with 100 nM dTAG V -1 ligand. 2 biological replicates at each time point are shown. Early apoptotic cells are defined as Annexin V + /DAPI- and late apoptotic cells as Annexin V + /DAPI+. D Western blot of apoptosis markers 24 h after treatment with 100 nM dTAG V -1 ligand. E Barplot of total % Annexin V positive cells quantified by flow cytometry after treatment with 100 nM dTAG V -1 ligand for 4 days in dTAG-TAK1 degron cells after knockout of indicated gene. F Barplot of competitive growth assay of dTAG-TAK1 cells expressing BFP and parental GSCs. Fold change of %BFP-positive cells in population after treatment with dTAG V -1 ligand for 7 days is shown relative to DMSO-treated control. G Cumulative growth assay in dTAG- TAK1 degron cells upon knockout of the second indicated gene by CRISPR and treatment with dTAG V -1 ligand. H Barplot depicting fold change of %BFP-positive dTAG-TAK1 cells in population after treatment with dTAG V -1 ligand for 7 days relative to DMSO-treated control and treatment with increasing concentrations of TNF ligand blocking antibody Etanercept. I Western blot of RIPK1 phosphorylation events after treatment with TNFα with or without TAK1 protein depletion. *denotes unspecific band. J Cartoon of molecular response to TAK1 inhibition in TAK1-dependent GSCs.

Article Snippet: For rescue experiments, cells were pre-treated for 1 h with 10 μM RIPK1 kinase inhibitor Necrostatin-2 (Nec-1s, Cayman Chemicals), 10 μg/ml Etanercept (Enbrel, Immunex Corp.) or 20 μM zVad-fmk (UBPBio), before addition of TAK1 inhibitor HS-276 or Takinib.

Techniques: Western Blot, Flow Cytometry, Knock-Out, Growth Assay, Expressing, Control, CRISPR, Blocking Assay, Phospho-proteomics, Inhibition

Journal: Cell Death & Disease

Article Title: TAK1 inhibition leads to RIPK1-dependent apoptosis in immune-activated cancers

doi: 10.1038/s41419-024-06654-1

Figure Lengend Snippet: A , B Barplot of %Annexin V positive cells ( A ) and fold cell expansion ( B ) of U3013MG cells with knockout of TNFR pathway members upon 4 days treatment with DMSO or 3 µM HS-276. C , D Barplot of %Annexin V positive cells ( C ) and fold cell expansion ( D ) of U3013MG cells treated for 4 days with HS-276 or Takinib in combination with RIPK1 inhibitor Necrostatin-1s (Nec-1s). E Cumulative growth assay of U3013MG or G166 cells treated with DMSO, HS-276, or a combination of HS-276 and Nec-1s. F Barplot of fold cell expansion within 4 days of treatment with DMSO or 3 µM HS-276 in 4 different glioma stem cell lines (U3013MG, G166, U3017MG, and G14). G Barplot of fold cell expansion within 4 days of treatment with DMSO or 3 µM HS-276 in fetal neural stem cells (fNSC, U5). H Western blot of death complex IIb formation in GSCs upon treatment with TNFα and HS-276 for 2 or 4 h. I Barplot of cell viability relative to DMSO in U3013MG cells treated with indicated chemotherapeutic drugs in increasing concentrations alone or in combination with HS-276 for 4 days.

Article Snippet: For rescue experiments, cells were pre-treated for 1 h with 10 μM RIPK1 kinase inhibitor Necrostatin-2 (Nec-1s, Cayman Chemicals), 10 μg/ml Etanercept (Enbrel, Immunex Corp.) or 20 μM zVad-fmk (UBPBio), before addition of TAK1 inhibitor HS-276 or Takinib.

Techniques: Knock-Out, Growth Assay, Western Blot